1. Field of the Invention
The present invention relates to a method for amplification of DNA from a blood sample, and a DNA amplification kit therefor, and more specifically to a DNA amplification method free of a need for setting a plurality of different process temperatures in a series of steps between a blood sample pretreatment step and a DNA amplification reaction step, and a DNA amplification kit therefor. The present invention is useful for life science researches and in medical diagnostic product-related fields.
2. Description of the Background Art
As a method for amplification of DNA from a blood sample, it is common to extract and purify genomic DNA and then subject the purified DNA to synthesis.
Lately, a SMAP (SMart Amplification Process) method has been being developed which includes the steps of: subjecting a blood sample to a pretreatment including no DNA purification treatment; and then amplifying DNA of the pretreated sample in an isothermal amplification system using a strand-displacing DNA polymerase to perform SNP typing.
The pretreatment step in the SMAP method includes performing heat denaturation at 98° C. for 3 minutes, using a 50 mM sodium hydroxide aqueous solution (see the following Non-Patent Document 1: Nature Methods, 2007, March; 4(3); 257-262).
[Non-Patent Document 1] Yasumasa Mitani, Alexander Lezhava, Yuki Kawai, Takeshi Kikuchi, Atsuko Oguchi-Katayama, Yasushi Kogo, Masayoshi Itoh, Toru Miyagi, Hideki Takakura, Kanako Hoshi, Chiaki Kato, Takahiro Arakawa, Kazuhiro Shibata, Kenji Fukui, Ryoji Masui, Seiki Kuramitsu, Kazuma Kiyotani, Alistair Chalk, Katsuhiko Tsunekawa, Masami Murakami, Tetsuya Kamataki, Takanori Oka, Hiroshi Shimada, Paul E Cizdziel & Yoshihide Hayashizaki, “Rapid SNP diagnostics using asymmetric isothermal amplification and a new mismatch suppression technology”, Nature Methods, 2007. March, Vol. 4, No. 3, pp 257-262.
The strand-displacing DNA polymerase has an optimum temperature at about 60° C. Therefore, in the SMAP method, the amplification step using the strand-displacing DNA polymerase can be performed under an isothermal condition (at about 60° C.) without temperature control of repeatedly increasing and reducing a process temperature as in a PCR (Polymerase Chain Reaction) method.
However, in the SMAP method, the blood sample pretreatment step to be performed in advance of the amplification step includes the heat treatment at 98° C.
This means that the SMAP method requires two types of process temperatures: a temperature for causing heat denaturation in the pretreatment step; and an optimum temperature for the strand-displacing DNA polymerase in the amplification step. Thus, it is necessary to provide two temperature control devices each operable to set a respective one of the two process temperatures, or a temperature control device, such as a thermal cycler, operable to change a process temperature.